Location: National Clonal Germplasm RepositoryTitle: Exogenous tocopherol and ascorbic acid improve in vitro recovery of cryopreserved Rubus shoot tips Author
Submitted to: In Vitro Cellular and Developmental Biology - Plants
Publication Type: Abstract Only
Publication Acceptance Date: 3/10/2009
Publication Date: 6/10/2009
Citation: Uchendu, E., Reed, B.M. 2009. Exogenous tocopherol and ascorbic acid improve in vitro recovery of cryopreserved Rubus shoot tips. In Vitro Cellular and Developmental Biology - Plants. 45:36. Interpretive Summary: Cold temperature stress can injure plants. Antioxidant compounds such as vitamins may reduce stress and prevent injury in cold environments. The objective of our study was to see if Vit E or Vit C could increase survival of plant parts subjected to liquid nitrogen temperatures (-320 F). We added the vitamins at 4 critical points in the cryogenic preservation procedure (pretreatment, loading, rinsing, recovery medium). Shoot tips of two blackberry cultivars were cryopreserved. Vit E and Vit C each improved recovery of shoots at several steps. Combinations of Vit E and Vit C produced survival that was similar to treatment with Vit C alone. Although treatments were effective at several critical points, we recommend adding Vit C to the pretreatment medium to produce the most effective growth response.
Technical Abstract: Oxidative processes involved in stresses such as cold temperatures can decrease the viability of plant tissues. Antioxidants that counteract these oxidative reactions could improve plant viability following the stresses involved in cryopreservation. We studied the effects of exogenous vitamin E (Vit E) and ascorbic acid (AA) added at four critical points of the cryopreservation process (pretreatment, loading, rinsing, recovery medium). Shoot tips of two blackberry cultivars were cold acclimated, then cryopreserved using a PVS2 vitrification protocol. Shoot tips (0.8 - 1 mm) of two Rubus hybrid cultivars were cold acclimated, then cryopreserved. Vit E added to the preculture medium for 48 h prior to liquid nitrogen exposure or to the rinse solution following rewarming significantly (P<0.0001) improved recovery of shoots compared to shoot tips without Vit E. There were no significant improvements with Vit E in the loading solution or the regrowth medium. Recovery of shoot tips treated with AA at three critical points was significantly better than untreated tissues. Recovery of shoot tips on standard regrowth medium with AA was significantly reduced compared to the recovery of untreated shoot tips. Regrowth medium without iron and with AA significantly improved regrowth compared to untreated shoot tips on either recovery medium. Combinations of Vit E and AA produced significant improvements in recovery at the four critical points compared with the untreated shoot tips, but the combination treatment was not significantly different from treatment with AA alone. Although treatments were effective at several critical points, we recommend adding AA to the pretreatment medium as the most convenient for the PVS2 vitrification process.